Optimizing HIV Protease Inhibitor Safety and Efficacy via Intracellular Targeting Abstract The AIDS epidemic is a global crisis with 31 million people worldwide who are living with HIV. According to the World Health Organization's 2010 revision of antiretroviral treatment therapy guidelines, creating less toxic therapies is a top priority to reduce adverse effects and improve compliance with therapeutic regimen. HIV protease inhibitors are a mainstay of highly active antiretroviral therapy (HAART) with annual sales of over $2.5 billion. All marketed HIV protease inhibitors (PI's) are substrates for cytochrome P450 and are co- administered with ritonavir, a pharmacokinetic booster. Boosting maintains therapeutic levels of compound to suppress viral replication and avoid incidence of viral mutation. Ritonavir is a strong inhibitor of the cytochrome P4503A4 isoform with a of Ki 5-70 nM which helps reduce P450 interactions of the co- administered PI. Eliminating P450 interactions of PI's would also eliminate the need for ritonavir. Toxicities attributed to ritonavir used alone or in combination with other HIV protease inhibitors (PI's) include hepatotoxicity, carotid artery thickening, hypercholesterolemia, hyperglycemia, and lipodystrophy. The elimination of P450 interactions for PI's is a high-priority research focus at several pharmaceutical companies, including Merck, Sequoia Pharmaceuticals, and Concert Pharmaceuticals. The elimination of ritonavir has been shown to reduce toxicities associated with HAART in short-term studies, and would increase compliance with therapeutic regimens. The specific aims of this proposal are: Aim 1. Design and synthesize a novel library of PI fragments linked to FKBP ligands. Aim 2. Screen and select library compounds for the ability to inhibit HIV protease employing an enzymatic assay and evaluate in vitro pk/pd. Aim 3. Re-screen the best PI candidates resulting from Aim 2 for microsomal stability, potency in cell infectivity assays, and p-glycoprotein assays to select candidates for further preclinical evaluation. PUBLIC HEALTH RELEVANCE: HIV patients who fail to comply with HAART have at least a 7-fold increased risk of mortality per patient year compared with HAART adherent patients. A recent meta-analysis of data showed adherence rates of only 55% in North America. HAART-associated toxicities are a major cause of non-adherence with as many as 37% of patient requiring modifications to their treatment due to adverse events over a two year period. By reducing the toxicity of HIV protease inhibitors, we hope to improve compliance with HAART and therapeutic outcomes.